DE60034029T2 - METHOD FOR PRODUCING SOLID PHASE MATRICES FOR RAMANSPEKTROSCOPY UNDER REINFORCING SURFACE APPLICATION (SERS) - Google Patents

Abstract

A method of forming a solid matrix for use with surface-enhanced Raman spectroscopy is described. The method comprises the steps of: admixing a colloidal metal solution with a polymeric support medium to form a suspension; optionally depositing said suspension on a surface; and then drying the suspension to form the matrix. The polymeric support medium provides a plymer/sol suspension in which the sol particles are resistant to aggregation and precipitation. Upon drying the suspension shrinks to provide a mechanically-hard film subsequently usable to provide a sample for spectroscopic analysis.

Description

The The present invention relates to a solid matrix for use with surface-enhanced Raman spectroscopy and to a method of making such a matrix.

spontaneous Raman scattering of chemical compounds is inherent weak effect; only a very small fraction of all photons on a normal nonabsorbent sample are Raman-scattered. By using the "Resonance Raman Effect" where the wavelength of the Light source selected is going to be on or near an electronic bandwidth of the sample in which light absorbs is able to fall increased Scattering probabilities are induced. However, the resonance Raman effects can only for Samples are used where the compound is of concern electronic bandwidth in which light is absorbed, at one the excitation wavelengths, the available to the experimenter standing, has.

One second method of increasing The signals of Raman scattering is the use of "surface enhancement" in so-called "surface enhanced Raman spectroscopy (SERS) "Under conditions where both the surface as well as the resonance gain The technique is called "surface-enhanced resonance Raman spectroscopy (SERRS) ". The surface reinforcement of Raman signals will observed when the kind of concern on or near one microscopically rough metal surface is absorbed. Not all metals let the effect come up; the two metals, the most common are used are silver and gold. A very wide range a method has been used to treat these metals, around surfaces to provide the Raman signals of chemical compounds (i.e. which are SER (R) S-active). The ones most often are those in which electrochemical or chemical Roughening of metal surfaces, the deposition of the metal on the substrate (for example, by the preparation of metal island films) and the preparation of colloidal Suspensions of metals, usually in aqueous solutions, and hereafter as sols designated, is involved.

• 1 Die Nichtreproduzierbarkeit der Präsentation der Probe an das optische System der Raman-Anregung/Raman-Sammlung. Dies ist ein rein mechanisches Problem und wird hier nicht weiter betrachtet.
• 2 Die Nichtreproduzierbarkeit in den SER(R)S-aktiven Oberflächen. Dies ist ein wesentliches Problem. Falls geraute Elektroden verwendet werden, muss der Aufrauungsvorgang zwischen den Messungen exakt wiederholt werden, und selbst wenn dies möglich ist, ist es schwierig, die Verunreinigung der Oberfläche durch hoch streuende Verbindungen zu eliminieren. Das Problem der Verunreinigung kann beseitigt werden, falls für jede Messung eine komplett frische Oberfläche verwendet wird. Die einfachste Art und Weise, frische Oberflächen für die Messung sicherzustellen, ist, kleine aliquote Teile von kolloidalen Lösungen (die kostengünstig bereitet werden können) zu verwenden und diese dann zu verwerfen. Es ist jedoch weithin anerkannt, dass die Bereitung von kolloidalen Lösungen mit identischen oberflächenverstärkenden Eigenschaften äußerst schwierig ist. Darüber hinaus sind die Lösungen von Natur aus instabil und können mit der Zeit oder aufgrund des Vorhandenseins von Spuren chemischer Unreinheiten zerfallen.

SER (R) S can be a very sensitive technique (lately, evidence of single molecules has been published) and has good delineation since it produces vibrational spectra of compounds characteristic of each compound. The combination of sensitivity and demarcation makes SER (R) S an obvious technique for analyzing a very wide range of chemical substances. Although the potential of the technique is clear, there has been very little utilization of the technique for routine analytical tasks. The main obstacle to routine analysis is that of signal reproducibility. There are two sources of this non-reproducibility:

• 1 The non-reproducibility of the presentation of the sample to the optical system of the Raman excitation / Raman collection. This is a purely mechanical problem and will not be considered here.
• 2 Nonreproducibility in the SER (R) S-active surfaces. This is a significant problem. If roughened electrodes are used, the roughening process must be exactly repeated between the measurements and, if possible, it is difficult to eliminate the contamination of the surface by highly scattering compounds. The problem of contamination can be eliminated if a completely fresh surface is used for each measurement. The simplest way to ensure fresh surfaces for the measurement is to use small aliquots of colloidal solutions (which can be prepared inexpensively) and then discard them. However, it is widely recognized that the preparation of colloidal solutions having identical surface-enhancing properties is extremely difficult. In addition, the solutions are inherently unstable and may degrade over time or due to the presence of traces of chemical impurities.

WO9724297 discloses that a mixed ceramic intermediate comprising a mixture of the Ceramic polymer and the nanoscale metal particles, can be prepared by a stable colloidal dispersion of Metal particles in an organic solvent solution of Ceramic precursor Poylmers or oligomer which serves to stabilize the particles, followed by evaporation of the solvent.

US5609907 discloses the formation of a self-assembled monomolecular layer of metal colloid metal particles on a polymer immobilized substrate for SERS bio-detection, in which a polymer-derivatized substrate, e.g. B. is formed from a macroscopic glass disc coated with polymerized 3-aminopropyltrimethoxysilane (APTMS) is immersed in a solution of colloidal metal particles (eg., Au, Ag). The metal particles have an affinity for certain organic functional groups of the polymerized organic films, so that the particles are immobilized at such locations, after which the film is dried.

An object of the present invention is to provide low cost of production of colloidal suspensions capable of producing a large number of identical and stable SER (R) S-active materials. In addition, these materials should be presented in shapes that are convenient to handle and manipulate, but inexpensive enough to be used once and then discarded.

According to the present The invention relates to methods of forming a solid matrix for use provided with surface enhanced Raman spectroscopy, as in the claims is defined.

The polymeric carrier medium surrounds the particles in the colloidal metal solution (sol), which in fact gives a polymer / sol suspension. If in this form, are the Sol particles are resistant to aggregation and deposition but still for each in a solvent supported analytes. In fact, it has been found that such suspensions are stable, insofar as they are over no perceptible spectral changes for several months demonstrate.

Of the Term "suspension" as used herein means any solid and / or liquid Form of the combination of the components, including gels and emulsions.

Complete drying leaves one mechanically hard, transparent or translucent film back. In the dry films, the metal particles are not only from it they are also protected against environmental damage. The Movies show no perceptible change after several months of storage. An inert matrix is made of metal particles and dry polymeric carrier medium has been formed, although between these substances no active binding has been formed. Such a matrix may then serve as a support surface for Analysis by SER (R) S can be used.

Around To activate the solid matrix (new), you can then use a suitable one solvent be treated, or even more common, a solution from analytes, at which time the matrix is due to the Penetration of the solvent swells and the particles contained in it are then free again, to interact with the chemical substance to be analyzed. Most importantly, the matrix regains its ability to enhance the surface produce after being solvated again.

In Any suitable polymeric carrier medium may be used in the present invention be used that capable is to form a suspension with sols suitable for use in SER (R) S and their properties of surface reinforcement are not reduced. In general, sols are due to the very small size Particles therein called "unstable", the therefore very sensitive to any change, especially any change in the environment, are. The polymeric carrier medium poses both in every liquid Shape or in any solid form a support for the sol-metal particles ready.

suitable polymeric carrier media include all known absorbents or hydrophilic swelling Polymers, such as those with carboxyl side chains, including polymers such as polycarbophil, copolymers such as hydroxyethyl methacrylate with methacrylic acid ("HEMA"), polyvinylmethyl-maleic anhydride ester and cellulose-based substances such as hydroxyethyl cellulose.

The polymeric carrier medium can be solid or liquid be.

Where the polymeric carrier medium entirely or essentially a liquid or a gel, etc., d. H. the formed suspension is at least not Solid, the suspension is applied to a surface or on a surface surface deposited, z. B. over a carrier spread, and then dried in air or under a vacuum, etc. During the Dry shrink the polymer / sol suspension as the polymeric carrier medium returns to an anhydrous state.

The Surfaces, to which the polymer / sol suspension can be applied every shape, design or shape. A conventional form is one in general made of clear glass flat plate. Alternatively, the suspension on multiple plates or holes applied, for. B. a (standard) micro-plate. The suspension could z. B. also be added to the inside of capillary tubes or pipettes, the tubes or pipettes can be used to prepare very small samples of analyte directly.

Preferably shows every liquid Suspend at least some viscosity on to their application a surface to facilitate. A gel suspension can be generally easy on a surface forming a screen.

Where the polymeric carrier medium wholly or substantially a solid, z. For example, as a crosslinked carbon-hydrophilic polymer or copolymer, such as HEMA, the colloidal metal solution may be added directly thereto to form the suspension. The carrier medium can aufquillen is However, return to its design, as soon as the suspension z. B. in air or under vacuum, etc. is dried. Such a carrier medium could be provided in the form of a sheet which could be divided into a plurality of suitable matrices (each one for subsequent re-feasible analysis) once the colloidal metal solution is absorbed or suspended thereby.

While everyone Metal particles used in Raman spectroscopy could in the colloidal metal solution of the present invention are such metal particles generally either silver or gold.

in the Generally, the solid matrix is a flat sheet or film or thin film is on a flat surface, such as a plate, or on a single or multiple perforated plate.

The The present invention clearly demonstrates the ability to produce a number to prepare identical and stable SER (R) S-active support surfaces, which have a can be stored for a relatively long period of time and which is an active medium can provide for use at any time and whose variation should be sufficiently minimal to be of the same Analytes thoroughly or essentially similar To provide Raman spectra. Consequently, now is a reproducibility of spectra over much longer Time periods than previously possible, and there should be much less time and effort involved in trying each time a Raman spectrum is desired, the analyte carrier conditions to repeat.

It It is contemplated that the present invention will be made possible could, that the use of Raman spectroscopy is much faster and broader fanned than previously used, e.g. B. outside forensich laboratories and much closer at the source of analytes. Their local use in forensic and / or medical diagnostics is clearly possible.

embodiments The present invention will now be described by way of example only with reference to the accompanying drawings, in where;

1a Figure 3 is a schematic drawing of two methods of forming a solid matrix according to the present invention.

1b is an assembly of standardized sampling in multi-well plates. Each hole in the plate contains a dried polymer / sol matrix according to the present invention which is activated by the addition of the analyte in the solution.

2 Figure 3 shows the surface enhanced Raman spectra of the putative anti-cancer drug AQ4N taken at two different excitation wavelengths in Noveon / Silver films.

3 shows the surface-enhanced Raman spectra of the putative anti-cancer drug AQ4N taken over a 500x concentration range in Noveon / Silver films.

4 shows the surface-enhanced Raman spectra of the putative anti-cancer drug AQ4N taken in a Gantrez / Silver film.

5 Figure 8 shows the repetition measurements of the surface enhanced Raman spectra of the putative anti-cancer drug AQ4N taken from eight different Noveon / Silver films. The internal standard was crystal violet.

6 - 11 For example, the surface enhanced Raman spectra (recorded in Noveon / Silver films) show a range of chemically distinct compounds, the range of compounds to which this invention can be applied and the range of excitation wavelengths that have already been used.

Materials and procedures

Aqueous silver and gold sols, as in 1a were prepared according to the procedures of the standard literature and were either used directly or concentrated by centrifugation using previously mentioned methods.

In one arrangement, by adding one of the liquid hydrophilic swelling polymers (see below) to the aqueous sol, viscous polymer / sol solutions were prepared as in the left part of 1a is shown. The mixture was allowed to stand for several hours with occasional stirring to allow hydration of the polymer prior to a final thorough mixing phase. The viscosity of the polymer / sol solutions can be varied simply by changing the concentration of the polymer in the solution.

The liquid or gelatinous polymer / sol solution was then coated on one of a number of different substrates, including metal, plastic, glass, and quartz supports (e.g., flat glass plate, as in Figs 1a shown), the holes of micro-well plates (as in 1b shown), the inner surfaces of hollow capillary tubes and deposited on the tips of optical fibers. The Deposited layers were then dried to a film (drying times depending on the thickness of the layer, ranging from 2 hours, for layers <100 μm thick, to two days, for layers that were - 5 mm thick). The dried films were stored in a dry environment. Alternatively, after drying, the films could be scraped from the backing and then pulverized. When this powder was redissolved it formed a SER (R) S active solution.

In a different arrangement and as in the right part of 1a For example, the sol was added directly to a solid, hydrophilic, swelling polymer, such as HEMA, to form a solid suspension. This was dried to give a solid matrix that was also stored in a dry environment before use. When this matrix was contacted with an aqueous analyte solution, it was found to be SER (R) S-active.

The solid hydrophilic swelling polymer could be either before or after Add The sols will be shared in order to be reusable Performance from the same sol a number of wholly or substantially to provide identical solid matrices. On similar Way could from a liquid Suspension a number of deposits are produced at the same time a To provide number of matrices or solid formed thereby Matrix could subsequently be divided. Consequently, you can with ease large numbers are formed on identical and stable solid matrices.

The Raman spectra were recorded using excitation at 458, 488, 514, 633, or 785 nm (Spectra-Physics 2020 Ar ⁺ laser or Spectra-Physics Ti / sapphire laser pumped from a Spectra-Pysics 2020 Ar ⁺ - Laser, typically 10-100 mW on the sample), using a 180 ° backscatter geometry. Scattered light was collected, passed through a holographic notch filter from Kaiser Optical System and then dispersed through a single stage (single stage) HR640 spectrogram from Jobin-Yvon onto a liquid N ₂ LN1152 CCD detector from Princeton Instruments. The spectra were typically accumulated for 120 seconds and were exported to the Spectral Manipulation Program Pack "LabCalc" for processing and presentation The spectrometer was calibrated using toluene and acetonitrile using a standard 50/50 mixture.

Results

The Raman signals derived from the polymer / sol matrices (either before or after drying / rehydration) were comparable intensity with those obtained from the simple sols to those added the same analytes had been. The Raman signals due to the polymer carrier medium There was negligible interference and there was no significant increase in Hintergrundlumineszenzintensität. The rehydrated films acted in exactly the same way like the more traditional ones Sols from which they had been prepared. There is no reason to believe that the range of analytes by conventional SER (R) S can not be studied, even with the use of Polymer / sol systems described herein can.

Around To demonstrate this point, signals have become broad Cross section of chemical compounds over the wavelength range 457.9-785 nm recorded. To record copies of each class on connections, studied using the polymer / sol combinations could become, or even the enormous range of compounds that use conventional SER (R) S has already been studied to duplicate, would be one enormous task. The spectra of a number of samples passing through the SER (R) S-active Reinforced films have been described above, but are hereby shown. The samples span porphyrins, azo dyes, indicators, metal complexes and therapeutic drugs. Shown are the spectra of the Center and two extremes of the wavelength range being studied has been.

The Spectra hereby were mainly used by get the movies over a liquid Prepared with a crosslinked polyacrylic acid (polycarbophil, Trade name Noveon AA1 by B.F. Goodrich), however, too an example is shown that of a polyvinylmethyl-maleic anhydride (Trade name Gantrez, I.S. P. Ltd.). There are many Polymers available under various trade names similar to those above. They are used in pharmaceutical products, cosmetic products and Cleaning products extensively used. The polymers used in the Trade available (even a single manufacturer can make a complete series produce) differ in rheological properties, because they are different networked, or slightly different Have main chains of the polymer.

The spectra herewith also show data taken from a single specimen, the supposed anticancer drug AQ4N, over a wide concentration range and at two different excitation wavelengths. The reproducibility of the films that formed are also shown. Shown herein are repeated spectra of a sample of AQ4N, the spectra being from small aliquots of the sample added to different dehydrated films. The spectra were normalized by adding an internal standard. Under these conditions, even a rough quantitative analysis (rationing peak heights of the strongest band in the sample and the standard) shows only a very small 3% deviation over the sample set taken.

Claims (12)

A method of forming a solid matrix for Use with surface-enhanced Raman spectroscopy, which includes the following steps: Blend a colloidal metal solution with a polymeric carrier medium, to form a suspension; and Drying the suspension, to form the matrix; the suspension before drying on a surface is deposited and the surface a microhole plate or the surface is the inside of a capillary tube or a pipette. Method according to claim 1, wherein a viscous suspension is formed. A method of forming a solid matrix for use with surface-enhanced Raman spectroscopy, which includes the following steps: Blend a colloidal metal solution with a polymeric carrier medium, to form a suspension; and Drying the suspension, to form the matrix; wherein the polymeric carrier medium a thin film or a thin one Location is. Method according to one the claims 1 to 3, wherein the polymeric carrier medium is an absorbent. Method according to one of the preceding claims, wherein the polymeric carrier medium is a hydrophilic polymer with carboxyl side chains. Method according to one of the preceding claims, the surface completely or is formed essentially of a material that is made of the Group consisting of glass, plastic, metal and quartz, is selected. Method according to one of the preceding claims, wherein the polymeric carrier medium Polycarbophil or polyvinylmethyl-maleic anhydride ester. Method according to one the claims 1 to 6, wherein the polymeric carrier medium is a cellulose-based substance. Method according to claim 8, wherein the cellulose-based substance is hydroxyethylcellulose is. Method according to claim 3, wherein the polymeric carrier medium a copolymer of hydroxyethyl methacrylate and methacrylic acid. Method according to one of the preceding claims, the metal being in the metal solution Silver or gold is. Method according to one the previous claims, the solid matrix is subsequently pulverized becomes.